PROJECT SUMMARY/ABSTRACT Monocytes and macrophages (M?) sense the presence of pathogens, tissue damage, and host-derived mediators in their environment and respond by differentiating into distinct functional phenotypes that mediate host innate immune responses. The process of M? differentiation has often been described in terms of ?plastic- ity,? implying that these cells modulate their functions through rapidly reversible differentiation steps in re- sponse to environmental changes in the host. For example, ?classically activated? M? (CAM; M1) are highly microbicidal, yet their production of inflammatory cytokines and nitric oxide may also damage host tissue. At the other end of the functional spectrum, ?alternatively activated? M? (AAM or M2), induced by IL-4 and IL-13, mediate ?wound healing? through elimination of damaged tissue and other anti-inflammatory mechanisms. Dur- ing the past three decades, the PI has undertaken research designed to dissect the complex molecular under- pinnings of M? differentiation and how this impacts host defenses and disease outcome. In the proposed stud- ies, the central hypothesis to be tested is that activation of specific intracellular signaling pathways distal to en- gagement of TLR4 and/or other signaling receptors by influenza-induced PAMPs and DAMPs serve to pro- gram expression of discrete cassettes of pro- and anti-inflammatory genes that control the manner in which the host M? responds to infection. Two Specific Aims are proposed to test this hypothesis in vitro and in vivo, with the ultimate goal of identifying novel therapeutic interventions for diseases where M? are required for contain- ing the invading pathogen and/or resolving tissue damage caused by pathogens or the host inflammatory re- sponse to infection. Both male and female mice (because of the availability of genetically engineered strains), and cotton rats (that are uniquely susceptible to human non-adapted isolates of influenza) will be utilized as models of primary (1o) influenza infection and 1o influenza infection followed by secondary (2o) bacterial infec- tion. The proposed innovative experimental approaches are designed to: (1) identify TLR4-interacting signaling receptors (i.e., PAR2, RAGE, CD11b/CD18) and delineate the contributions of these interactions to regulation of M? differentiation/susceptibility to influenza infection; and, (2) identify influenza-triggered, epigenetic and/or M? differentiative mechanisms that mediate susceptibility to 2o Gram-positive bacterial infection. At the conclu- sion of these comprehensive studies, key processes that govern interactions of TLR agonists that lead to changes in M? activation will have been defined that, in turn, can be expected to translate into reasonable and practical therapeutic approaches for controlling invading pathogens or counteracting inflammatory damage to tissues induced by pathogens, ultimately providing evidence-based therapies to treat infectious diseases.